The long-term goal of this research proposal is to determine at the molecular level the functional role of microfilaments in the processes of nerve growth and axoplasmic transport. Actin will be isolated from chick brain in high yield and its various microheterogeneous forms will be separated and characterized by amino acid analysis and peptide mapping. The amount and distribution of actin will be determined by polyacrylamide gel electrophoresis within neurons, astrocytes and glial cells separated from whole brain. Optimal conditions for brain actin assembly will be determined with regards to pH, ionic strength, nucleotide requirements, monovalent and divalent metal ion requirements, and the stoichiometry of the heterogeneous actin species within actin oligomers. Spectroscopic studies using UV absorbance and circular dichroism measurements will be used to study the conformation of brain and muscle actin in solution and to follow conformational changes which occur in the assembly of actin filaments. The effects of other brain proteins on the assembly process will also be investigated. The mechanism by which local anaesthetics inhibit nerve growth by disruption of functioning microfilament system will be investigated. Using electron microscopy and spectroscopy methods for studying both the assembly of brain microfilaments and their interaction with other brain proteins in vitro, the effects of anaesthetics on these processes will be measured. Comparison of these effects with those caused by mono and divalent metal ions should distinguish direct effect of anathetics on these processes from membrane mediated events.